Whole genome doubling drives oncogenic loss of chromatin segregation [WGS]

Whole genome doubling (WGD) is a recurrent event in human cancers and it promotes chromosomal instability and acquisition of aneuploidies. However, the 3D organization of the chromatin in WGD cells and its contribution to oncogenic phenotypes are currently unknown. Here, we show that in p53 deficient cells WGD induces loss of chromatin segregation (LCS), characterized by reduced segregation between short and long chromosomes, A and B sub-compartments, and adjacent chromatin domains. LCS is driven by downregulation of CTCF and H3K9me3 in cells that bypassed activation of the tetraploid checkpoint. Longitudinal analyses revealed that LCS primed genomic regions for sub-compartment repositioning in WGD cells, which resulted in chromatin and epigenetic changes associated with oncogene activation in tumours ensuing from WGD cells. Importantly, sub-compartment repositioning events were largely independent of chromosomal alterations, indicating that these were complementary mechanisms contributing to tumour development and progression. Overall, LCS initiates chromatin conformation changes that ultimately result in oncogenic epigenetic and transcriptional modifications, suggesting that chromatin evolution is a hallmark of WGD-driven cancer. For mutational and copy number change analyses in post-WGD cells in RPE TP53-/- line, whole genome sequencing was performed in RPE TP53-/- control, WGD, and 6w and 20w pWGD cells grown in vitro and in vivo. Biological replicates for 6w pWGD were considered and are denoted as experiment 1-4 based on multiple WGD inductions. RPE cells with a TP53 wild type background were considered as reference for calling mutations and copy number changes.

全基因组加倍（Whole genome doubling, WGD）是人类癌症中的复发性事件，可促进染色体不稳定及非整倍体的获得。然而，目前学界对于全基因组加倍细胞内染色质的三维组织结构，及其在致癌表型形成中的作用仍不明晰。本研究发现，在p53缺陷型细胞中，全基因组加倍会诱导染色质分离丧失（Loss of chromatin segregation, LCS），其特征为长短染色体、A与B亚区室以及相邻染色质结构域之间的分离程度降低。LCS的发生由绕过四倍体检查点激活的细胞中CCCTC结合因子（CTCF）与组蛋白H3第9位赖氨酸三甲基化（H3K9me3）的表达下调所驱动。纵向追踪分析显示，LCS可使全基因组加倍细胞中的基因组区域具备亚区室重定位的潜能，最终导致继发于全基因组加倍细胞的肿瘤中出现与癌基因激活相关的染色质及表观遗传改变。值得注意的是，亚区室重定位事件在很大程度上独立于染色体改变，这表明二者是促进肿瘤发生与进展的互补机制。综上，LCS可启动染色质构象改变，最终引发致癌性的表观遗传与转录组修饰，这提示染色质演化是全基因组加倍驱动型癌症的标志性特征。为分析视网膜色素上皮（Retinal pigment epithelium, RPE）TP53基因敲除细胞系中全基因组加倍后细胞的突变与拷贝数变化，本研究对体外及体内培养的RPE TP53敲除对照细胞、全基因组加倍细胞，以及培养6周与20周的全基因组加倍后细胞开展了全基因组测序。本研究纳入了6周龄全基因组加倍后细胞的生物学重复样本，并基于多次全基因组加倍诱导实验将其标记为实验1至实验4。本研究以TP53野生型背景的RPE细胞作为突变与拷贝数变异检测的参考样本。